I. Tumor Suppressor Genes
Cancer is a set of diseases resulting from uncontrolled cell growth, which causes intractable pain and death for more than 300,000 people per year in the United States alone. Oncogenes and tumor suppressor genes are at opposite ends of a spectrum of gene actions that either promote or retard cancer cell growth. The development of cancer is believed to depend on the activation of oncogenes and the coincident inactivation of growth suppressor genes (Park, M., “Oncogenes” in The Genetic Basis of Human Cancer (B. Vogelstein et al., eds.) pp. 205-228 (1998)). Oncogenes are mutated, dominant forms of cellular proto-oncogenes that stimulate cell proliferation, while tumor suppressor genes are recessive and normally inhibit cell proliferation (Cooper, 1995). The loss or inactivation of tumor suppressor genes is widely thought to be one of the contributors to unregulated cancer cell growth. While the discovery and identification of oncogenes has been relatively straightforward, identifying tumor suppressor genes has been much less so (Fearon, The Genetic Basis of Human Cancer (B. Vogelstein et al., eds.) pp. 229-236 (1998)).
Both oncogenes and tumor-suppressing genes have a basic distinguishing feature. The oncogenes identified thus far have arisen only in somatic cells and thus have been incapable of transmitting their effects to the germ line of the host animal. In contrast, mutations in tumor-suppressing genes can be identified in germ line cells and are thus transmissible to an animal's progeny. About a dozen such tumor suppressor genes have been identified, with the hope that knowledge of their mechanism(s) might yield therapeutically relevant insights.
Tumor suppressor gene action depends on either mutation or deletion of both tumor suppressor alleles or on a reduction in the absolute level of expressed tumor suppressor protein. In their natural state, tumor suppressor genes act to suppress cell proliferation. Damage in such genes leads to a loss of this suppression, and thereby results in tumorigenesis. Knudson's “two-mutation hypothesis” is a well studied statistical model for tumor suppressor gene action which is based on the epidemiological analysis of retinoblastoma. (Knudson, A. G., Proc. Nat. Acad. Sci. USA. 68:820-823 (1971)). According to this model, the host is heterozygous for the tumor suppressor gene, and cancer ensues when the single remaining functional allele also mutates to create a nullizygous state. An alternative model is the “haplo-insufficient hypothesis” in which the tumor cell produces abnormally low levels of wild type tumor suppressor gene product. Thus, in both of these models the deregulation of cell growth may be mediated by the inactivation of tumor-suppressing genes (Weinberg, R. A., Scientific Amer., September 1988, pp 44-51).
Tumor suppressor genes are principally known for control of cell proliferation by their action on the cell cycle. Well-studied examples include Rb (Weinberg, R. A., Cell, 81:323-330 (1996)), p53 (Greenblatt, M. S. et al., Cancer Res. 54:4855-4878, (1994); Williams, B. O. et al., Cold Spring Harbor Symp. Quant. Biol. 59:449, (1994)); Levine, A. J., Cell 88:323-331 (1997)), and p16 (Cairns, P., et al., Nat. Genetics 11:210-212, (1995)); Okamoto, A., et al., Cancer Research 55:1448-1451, (1995)). Another example of a tumor suppressor gene acting on the cell cycle is the p27KIP1 gene, also known simply as p27, which physiologically inhibits cyclin-dependent kinases, and thereby blocks cell proliferation (Fero, M. L., et al., Nature 396:177-180 (1998)).
In understanding how tumor suppressor genes impact the cell cycle, one must understand that cell cycle transitions are regulated by specific cyclin dependent kinases that consist of an activating cyclin subunit and a catalytic Cdk subunit (Polyak, K., et al., Cell 78:59-66 (1994)); Hartwell, L., Cell 71:543-546, (1992)); Nurse, P., Nature 344:503-508,(1990)). The functions of the respective cyclins and Cdk's in mammalian cells correspond to the different phases of the cell cycle. For example, during the G1 phase, cyclin D-Cdk4/6 and cyclin E-Cdk2 are catalytically active and rate limiting for cell cycle progression. Growth factors induce the synthesis of D-type cyclins to initiate the G1 phase. The D-type cyclins then associate with Cdk4/Cdk6, and the active Cdk's then hyperphosphorylate Rb to drive the cell past the restriction point (Buchkovich, K., et al., Cell 58:1097-1105 (1989)); see Weinberg, R. A., Cell 81:323-330 (1996)). Tumor suppressor genes have been found to affect the function of both of these types of subunits.
In addition to the cell cycle, tumor suppressor genes can also control cellular differentiation by acting as transcription factors and/or by modulating specific downstream DNA repair targets involved in maintaining genomic integrity. In this class of tumor suppressor gene activity, inactivation of the tumor suppressor gene, p53, is the most common, resulting in a somatic mutation that causes malignancy (Nigro, J. M., et al., Nature 342:705-708 (1989); cf., review by Nguyen and Jameson, 1998). Of particular note, p53 is a frequent target for mutation in lung cancer (Takahashi, R., et al., Science 246:491-494 (1989)) and bladder cancers (Sidransky, D., et al., Science 252:706-709 (1991)). A germline mutation for p53 is the basis for a familial cancer, the Li-Fraumeni syndrome (Srivastava, S., et al., Nature 348:747-749 (1990)). At the level of DNA repair, p53 works in the following manner: When DNA is damaged, a resulting signal causes stabilization of p53, which in turn causes transcriptional deregulation of p21, resulting in cell cycle arrest in the G1 phase (Hunter, T., Cell 75:839-841 (1993)).
Finally, tumor suppressor genes have also been implicated in controlling apoptotic cell death (Graeber, T. G., et al., Nature 379:88 (1996)). Again, p53 figures prominently in this process as well (Basu, A., et al., Mol. Hum. Reprod. 4:1099-1109 (1998)). The clear message from this brief summary is that the individual tumor suppressor genes cannot be viewed from a single perspective.
In order to study these tumor suppressor genes, model systems must be developed. Recent advances in recombinant DNA and genetic technologies have made it possible to discover and assess new tumor suppressor genes. One of the key model systems available is the transgenic animal. Such animals have been engineered to contain gene sequences that are not normally or naturally present in an unaltered animal. The techniques have also been used to produce animals which exhibit altered expression of naturally present gene sequences.